Carbachol triggers RyR-dependent Ca2+ release via activation of IP3 receptors in isolated rat gastric myocytes
Carbachol triggers RyR-dependent Ca2+ release via activation of IP3 receptors in isolated rat gastric myocytes
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DOI：
10.1113/jphysiol.2002.020131
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发表时间：
2002-08-01
影响因子：
5.5
通讯作者：
McGeown, JG
中科院分区：
文献类型：
--
作者：
White, C;McGeown, JG
在离体大鼠胃肌细胞中，使用激动剂诱发的Ca ~（2+）信号研究了不同细胞内Ca ~（2+）释放通道之间可能的相互作用。自发的，局部Ca 2+瞬变，观察在荧光-4-负载细胞与线扫描共聚焦成像。这些被阻止ryanodine（100妈妈），但不是由肌醇1，4，5-三磷酸受体（IP 3R）阻滞剂，2-氨基乙氧基二苯基硼酸酯（100妈妈），确定他们作为钙火花。咖啡因（10毫米）和卡巴胆碱（10妈妈）启动Ca 2+释放的网站，共同定位与对方和任何Ca 2+火花网站。在fura-2负载的细胞中，细胞外2-氨基乙氧基二苯基硼酸盐和细胞内肝素（5 mg ml（-1））均抑制卡巴胆碱引起的整体细胞质[Ca 2 +]瞬变，证实它是IP 3R依赖性的。2-氨基乙氧基二苯基硼酸酯和肝素也增加了对咖啡因的反应。这可能反映了增加的Ca 2+存储内容，因为2-氨基乙氧基二苯基硼酸盐离子霉素诱发的瞬变幅度增加了一倍以上。Ryanodine完全消除卡巴胆碱和咖啡因的反应，但只减少离子霉素瞬变30%，这表明Ryanodine对卡巴胆碱瞬变的阻断不仅仅是由于存储耗尽。IP（3）R和RyR的双标记显示它们的分布有广泛的重叠。这些结果表明卡巴胆碱通过IP（3）Rs和RyRs之间的协同作用刺激Ca ~（2+）释放，IP（3）Rs参与Ca ~（2+）库含量的调节。
Possible interactions between different intracellular Ca2+ release channels were studied in isolated rat gastric myocytes using agonist-evoked Ca2+ signals. Spontaneous, local Ca2+ transients were observed in fluo-4-loaded cells with linescan confocal imaging. These were blocked by ryanodine (100 mum) but not by the inositol 1,4,5-trisphosphate receptor (IP3R) blocker, 2-aminoethoxydiphenyl borate (100 mum), identifying them as Ca2+ sparks. Caffeine (10 mm) and carbachol (10 mum) initiated Ca2+ release at sites which co-localized with each other and with any Ca2+ spark sites. In fura-2-loaded cells extracellular 2-aminoethoxydiphenyl borate and intracellular heparin (5 mg ml(-1)) both inhibited the global cytoplasmic [Ca2+] transient evoked by carbachol, confirming that it was IP3R-dependent. 2-Aminoethoxydiphenyl borate and heparin also increased the response to caffeine. This probably reflected an increased Ca2+ store content since 2-aminoethoxydiphenyl borate more than doubled the amplitude of transients evoked by ionomycin. Ryanodine completely abolished carbachol and caffeine responses but only reduced ionomycin transients by 30%, suggesting that blockade of carbachol transients by ryanodine was not simply due to store depletion. Double labelling of IP(3)Rs and RyRs demonstrated extensive overlap in their distribution. These results suggest that carbachol stimulates Ca2+ release through co-operation between IP(3)Rs and RyRs, and implicate IP(3)Rs in the regulation of Ca2+ store content.